溶解
计算生物学
蛋白质组学
蛋白质-蛋白质相互作用
核糖体
枯草芽孢杆菌
功能(生物学)
结构生物学
质谱法
生物
细菌蛋白
化学
细胞生物学
细菌
生物化学
核糖核酸
遗传学
基因
色谱法
作者
Francis J. O’Reilly,Andrea Graziadei,Christian Forbrig,Rica Bremenkamp,Kristine Charles,Swantje Lenz,Christoph Elfmann,Lutz Fischer,Jörg Stülke,Juri Rappsilber
标识
DOI:10.1101/2022.07.26.501605
摘要
Summary Accurately modeling the structures of proteins and their complexes using artificial intelligence is revolutionizing molecular biology. Experimental data enables a candidate-based approach to systematically model novel protein assemblies. Here, we use a combination of in-cell crosslinking mass spectrometry, cofractionation mass spectrometry (CoFrac-MS) to identify protein-protein interactions in the model Gram-positive bacterium Bacillus subtilis . We show that crosslinking interactions prior to cell lysis reveals protein interactions that are often lost upon cell lysis. We predict the structures of these protein interactions and others in the Subti Wiki database with AlphaFold-Multimer and, after controlling for the false-positive rate of the predictions, we propose novel structural models of 153 dimeric and 14 trimeric protein assemblies. Crosslinking MS data independently validates the AlphaFold predictions and scoring. We report and validate novel interactors of central cellular machineries that include the ribosome, RNA polymerase and pyruvate dehydrogenase, assigning function to several uncharacterized proteins. Our approach uncovers protein-protein interactions inside intact cells, provides structural insight into their interaction interface, and is applicable to genetically intractable organisms, including pathogenic bacteria.
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